Obesity is a major health problem that predisposes individuals to diabetes, cardiovascular disease and cancer. Excessive caloric intake is linked to resistance of hypothalamic homeostatic centers to peripheral feedback signals and to hedonic processes involving limbic circuits that mediate the reinforcing value of palatable food. These neural pathways interact with each other, but the mechanisms are largely unknown. Our work has focused on hypothalamic proopiomelanocortin (POMC) neurons that, together with adjacent AGRP/NPY neurons, integrate interoceptive signals of nutritional state and regulate food intake and energy expenditure. We have developed a unique mouse model of hypothalamus-specific POMC-deficiency with the capacity for POMC re-expression. These mice exhibit genetically reversible hyperphagia, obesity, meal pattern alterations and decreased locomotor activity, and were used to test the functional importance of POMC neuron heterogeneity and the absolute number of POMC neurons required to regulate energy balance. We discovered that POMC neurons synaptically release GABA and glutamate and, by using a genetic strategy to fluorescently tag synaptophysin in POMC neurons, we identified discrete synaptophysin-positive boutons that were immuno- labeled either for POMC peptides, GABA, or glutamate. POMC neurons were shown to project to multiple brain areas that integrate energy status with motivational, rewarding and locomotor behaviors. Therefore, our goal for the next competing cycle of this project is to test the hypothesis that hypothalamic POMC neurons are in synaptic contact with limbic circuit neurons, physiologically modulate their activity through the differentil release of melanocortin peptides, ?-endorphin and amino acid neurotransmitters, and influence appetitive behaviors and locomotor activity related to food acquisition and consumption. We will experimentally address this overarching hypothesis through the following three specific aims. 1) Map the location and neurochemical properties of afferent and efferent neurons with monosynaptic connections to arcuate POMC neurons, focusing on the nucleus accumbens, bed nucleus of the stria terminalis, paraventricular nucleus of the thalamus, and the ventral tegmental area. 2) Directly measure basal levels and stimulated release of the neuropeptides and amino acid neurotransmitters produced by POMC neurons in the nuclei named above that comprise the limbic motivational circuit. These experiments will employ brain microdialysis directly coupled on-line to a novel capillary liquid chromatography/mass spectrometry system capable of simultaneously detecting multiple neuropeptides with picomolar sensitivity. 3) Test the hypothesis that POMC neuron/melanocortin projections to the mesocorticolimbic dopamine pathway modulate locomotor activity and appetitive motivation for food. Together, these investigations of hypothalamic POMC neurons will provide new mechanistic insights regarding the complex links between the homeostatic and hedonic control of feeding, dysregulation of which may underlie the obesity epidemic in the modern world.